The primary goal of our studies is to look for additional evidence for two alternative modes of priming during initiation of DNA replication forks: 1) by RNA priming from an origin sequence and 2) by DNA priming from the termini of recombinational intermediates. 1) We will test whether mutations which are singly defective is only one of the putative mechanisms and therefore are "leaky", become completely defective when they are combined in a double mutant. 2) We plan to view the vegetative DNA of these single and double mutants in the electron microscope and compare it to the wild type DNA which has been analyzed by R. Dannenberg in our lab. 3) Southern blots of intracellular T4 DNA will be extended, using different restriction enzymes and using DNA from different mutants and from wild type infections to determine to what extent certain mutations influence the decision between uni- or bidirectional replication and between different modes of initiation. 4) As so n as we have identified the origin region on a small restriction fragment we intend to sequence it. 5) We will continue to analyze the apparent instability of gene 32 which has been seen in many of our experiments during the last 5 years.